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Murine induced pluripotent stem cells (iPSCs) lacking IL-1 receptor type 1 (IL-1R1), engineered using the CRISPR/Cas9 system, are able to form cartilaginous tissue that is resistant to IL-1α-mediated inflammation, a new study shows. “We were able to effectively and efficiently create a new line of stem cells that did not previously exist in nature and were immune to the effects of cytokine-induced inflammation, but had no other detectable changes in their properties,” explains Farshid Guilak, one of the corresponding authors of the study.

Stem cell therapies hold great promise for tissue repair in inflammatory arthritis. However, the inflammatory microenvironment within arthritic or injured tissues remains a major problem, as it inhibits the differentiation capacity of stem cells and can degrade newly regenerated tissue. “Our goal was to apply the newly developed CRISPR/Cas9 method for gene editing to create a stem cell that does not respond to inflammatory cytokines,” says Guilak.

Stem cell therapies hold great promise for tissue repair in inflammatory arthritis

The researchers obtained 41 iPSC clones, four of which had a homozygous deletion of Il1r1(Il1r1−/−), and three that had a heterozygous Il1r1+/− genotype. Reduced or absent transcriptional activity of nuclear factor-κB (NF-κB) following stimulation with IL-1α confirmed the effective deletion of IL-1R1 in these clones.

In cartilage pellets derived from wild-type and Il1r1+/− iPSCs, treatment with IL-1α increased the expression of proinflammatory cytokines such as CC-chemokine ligand (CCL) 2 and IL-6, as well as extracellular matrix proteolytic enzymes such as ADAMTS4 and MMP9. By constrast, no significant differences were found in the expression of these inflammatory markers in cartilage derived from Il1r1−/− iPSCs. The investigators also found that IL-1α considerably reduced the levels of sulfated glycosaminoglycan in cartilage derived from wild-type and Il1r1+/− cells, whereas no such effect was seen in cartilage derived from Il1r1−/− cells.

These data suggest that CRISPR/Cas9 gene editing could be successfully used in iPSCs to generate cartilage that is resistant to cytokine-induced inflammation and degradation. “More advanced strategies could be developed that build on this proof-of-principle work, such as engineering the dynamic, autonomous control of cell responses to environmental stimuli or reprogramming epigenetic states,” remarks Charles Gersbach, co-corresponding author of the study.